Tatupu attended the University of Southern California, playing in the 1978 Hula Bowl and finished his career with 223 rushing attempts for 1277 yards (5.73 yards per attempt avg.). The Mosi Tatupu Award, given annually to the College Football Special Teams Player of the Year by the Maui Quarterback Club and the Hula Bowl, bears his name.

Drafted in 1978 by the New England Patriots (8th round, 215th overall), Tatupu played fullback and became an ace on special teams. He rushed the ball 612 times for 2,415 yards and 18 touchdowns; in the 1983 season he led the league with 5.5 rushing yards per attempt. He caught 96 passes for 843 yards and two touchdowns, against the Colts in 1981 and the Falcons in 1983. His season high for rushing yards came in 1983 with 578 rushing yards and four touchdowns, while in 1984 he set a personal best with 16 catches for 159 yards. In 1987 against the Raiders Tatupu threw a 15-yard touchdown to Tony Collins.

He was used mostly on special teams kick coverage. He did have seven kick returns for 56 yards in his career.

In The Simpsons episode "Treehouse of Horror III" Mosi Tatupu's name is used during the "King Homer" segment. This segment is a parody of the 1933 film King Kong. During this segment, the character of the chief of Ape Island is shown to say "Mosi Tatupu, Mosi Tatupu", which in their language means "the blue-haired woman will make a fine sacrifice". The blue-haired woman is Marge Simpson who is playing Fay Wray to Homer's King Kong. One of the most popular players to play for the New England Patriots, Tatupu even had his own section of fans at Foxboro Stadium, "Mosi's Mooses," who all adorned moose heads and continually chanted his name throughout every game he played at home.